Effects Of 4℃ Exposure On Growth,Estrous Cycle And Uterine Proteome Of Female C57BL/6 Mice

Objectives The purpose of this thesis is to study the effects of 4℃ exposure on growth,estrous cycle and uterine proteome of female C57BL/6 mice,which provides a theoretical basis for preventing reproductive harm caused by cold.Methods 1.Twenty eight-week-old SPF female C57BL/6 mice,within 17～20 g,were randomly divided into 2 groups,control group and cold group,10 in each group.Cold group was exposed to 4℃ in cold medical cabin for 14 days,4 hours per day,while control group stayed in common condition.Body weight and food intake were collected once two days.2.Vaginal smears were used to observe the estrous cycle.After 14 days,blood and organs were collected from each mouse after anesthetized and weighted.Serum levels of sex hormone and interleukins were determined by using mouse ELISA kits.The oxidative stress of uterus was determined.The uterus and ovary were prepared to observe the structural changes.3.Uteruses from control group and cold group,3 of each group,were detected by liquid chromatography-mass spectrometry(LC-MS)after hydrolyzed by tryspin.Differential proteins were found and quantified by Max Quant software.The R package cluster Profiler was used for Gene Ontology(GO)enrichment analysis.Protein-protein interactions(PPI)network was drawn by using STRING database.Results 1.Compared with the control group,4℃ exposure made the cold group urine and faeces more.Some mice lost hair,but apparent harm were not found.No mice died.2.Through entire experiment period,the 4th,6th,8th,and 10 th days of body weight showed that cold group was significantly higher than control group(P(27)0.05).Food utilization rate was not affected.The uterine weight and organ coefficient of the cold group were significantly lower(P(27)0.01).3.Compared with the control group,the diestrus phase of cold group was significantly increased after cold exposure(P(27)0.01)and it was(5.83?1.72)days.4.Compared with the control group,the cold group had significant higher serum level of FSH(P(27)0.01),(6.58?0.21)m IU/ml,and lower serum level of PRL(399.87?12.23)pg/ml and Prog(5.15?2.52)ng/ml(P(27)0.01).Serum level of E2 and LH showed no significance.5.Pathological examination showed that uterine glands were expanded and the amount of follicles was decreased in cold group.The cold group had significantly lower activities of uterus SOD(6.79?0.76)ng/ml(P(27)0.01)and higher content of uterus MDA(22.80?0.83)nmol/ml and CPR(245.31?13.45)ng/ml(P(27)0.01).6.71 genes were selected among these quantified genes by screening,in which 39 genes were up-regulated and 32 genes were down-regulated.Gene Ontology analysis results shows that in biological process,up-regulated differential proteins are mainly enriched to oxidative stress and vesicle-mediated transport in synapse,while downregulated differential proteins are mainly enriched to dicarboxylic acid metabolic process.In cellular components,up-regulated differential proteins are mostly enriched in vesicle,transport vesicle and synaptic vesicle,while down-regulated differential proteins are mainly enriched in polysome and cytosolic ribosome.In molecular function,up-regulated differential proteins are mainly enriched to polyubiquitin modificationdependent protein binding,modification-dependent protein binding,etc.,while downregulated differential proteins are mainly enriched to ribonucleoside binding,purine ribonucleoside binding,and purine nucleoside binding,etc.Protein-protein interactions shows that differential proteins caused by cold exposure can affect endoplasmic reticulum signal peptide transport.Conclusions 1.4℃ exposure can cause the body weight of female C57 mice to change in a short period of time,but it can not cause significant changes in food intake and food utilization.Cold stress could increase female C57BL/6 mice estrous cycle,especially diestrus.4℃ exposure could increase serum level of FSH and decrease serum level of PRL and PROG.2.4℃ exposure changes the expression levels of some proteins of uterus.The differential proteins are mainly related to oxidative stress,dicarboxylic acid metabolism,synaptic vesicles,and endoplasmic reticulum signal peptide transport.